Crystallization and preliminary crystallographic analysis of the first condensation domain of viomycin synthetase.

Nonribosomal peptide synthetases (NRPSs) are large multimodular enzymes that synthesize important secondary metabolites such as antibiotics. NRPSs follow a modular synthetic logic whereby each successive amino-acid monomer is added to the peptide chain by successive multi-domain modules. The condensation domain catalyzes the central chemical event in the synthetic cycle, peptide-bond formation, and is present in every elongation module of the NRPS. Viomycin is an antituberculosis nonribosomal peptide that is synthesized by a series of four NRPS proteins and then modified by tailoring proteins. In order to study the mechanisms of peptide-bond formation in viomycin and in NRPSs in general, a structural study of the first condensation domain of the viomycin synthetase protein VioA (VioA-C1) was initiated. The gene for VioA-C1 was cloned from genomic DNA of Streptomyces vinaceus, expressed as an octahistidine-tagged construct and purified by column chromatography. VioA-C1 was crystallized using the sitting-drop vapor-diffusion method. X-ray diffraction data were collected on a rotating-anode source to 2.9 Ã resolution. The data could be indexed in the orthorhombic space group P212121, with unit-cell parameters a = 46.165, b = 68.335, c = 146.423 Ã . There is likely to be one monomer in the asymmetric unit, giving a solvent content of 49.2% and a Matthews coefficient (VM) of 2.42 Ã (3) Da(-1). Structural determination is in progress.